Biofilms cost this nation billions of dollars yearly in equipment damage, product contamination, infections, and colonization of devices and prosthetic implants. The effectiveness of conventional methods of microbial eradication is drastically impaired with biofilms since they exhibit atypical characteristics when compared to their planktonic counterparts. Therefore the ability to destroy these organisms demands the development of alternative techniques. The broad, long term goal of this project is to fill a gap regarding the lack of effective biofilm inactivation/sterilization methods. The use of plasmas, the fourth state of the matter, offers a novel alternative since plasmas contain a mixture of reactive agents individually known as bacteria- killing agents. Although evidence suggests that plasma is useful to inactivate bacterial biofilms, there are still many unresolved questions before we can understand the mechanism that causes inactivation/sterilization and apply it for medical or environmental purposes. The main goal of this research is to understand the mechanisms leading to gas discharge plasma-assisted biofilm inactivation. The specific aims are: 1) to determine the effect of plasma on the physiological status of biofilms-forming cells. We will test the hypothesis that plasma affects the physiological status (viability, culturability, metabolic status) of biofilm- forming cells and the effect depends on the amount of time that biofilms have been exposed to plasma and the age of the biofilm. 2) to determine the effect of plasma on the structure biofilm-forming cells. We will test he hypothesis that plasma affects the structure (integrity of cell envelopes and eventual presence of pores) of biofilm-forming cells, and the effect depends on the amount of time that biofilms have been exposed to plasma and the age of the biofilm;and 3) to relate gas-plasma composition with biofilm inactivation/sterilization. We will determine whether the response of cells to plasma is a general phenomenon or if it depends on the type of plasma used. We will select a plasma environment with a particular diversity or abundance of reactive agents and we will use it to evaluate its effect on biofilm-forming cells structure and physiological status. This study will provide a comprehensive understanding of plasma-assisted-biofilm inactivation and its mechanisms, fulfilling NIH's goal #3 (to. expand the knowledge base in medical and associated sciences in order to enhance the Nation's economic well-being and ensure a continued high return on the public investment in research). By achieving this main goal we expect to build the basis for the future development of a technology that will, as a long-term goal, solve a serious medical problem, fulfilling NIH's mission (Science in pursuit of fundamental knowledge about the nature and behavior of living systems and the application of that knowledge to extend healthy life and reduce the burdens of illness and disability);and particularly goal # 1 (to foster fundamental creative discoveries , innovative research strategies, and their applications as a basis to advance significantly the Nation's capacity to protect and improve health). PUBLIC HEALTH RELEVANCE: Biofilms contaminate common household surfaces and many medical devices, (e.g., catheters, contact lenses, mechanical heart valves, joint replacements, etc.), causing illness and death. The huge doses of antimicrobials required to destroy biofilms are environmentally undesirable and may be hazardous to a patient. Therefore, the ability to destroy these organisms demands the development of alternative techniques. Plasmas, the fourth state of matter, offer an attractive alternative. This research will study the mechanisms of plasma-assisted- biofilm inactivation with the long term goal of developing an easy-to-use, non-hazardous sterilization method to destroy biofilms.